New Sensitive Method for the Measurement of Lysozyme and Lactoferrin for the Assessment of Innate Mucosal Immunity. Part I: Time-Resolved Immunofluorometric Assay in Serum and Mucosal Secretions

Abstract

Mucous peristalsis, mucus and immunity proteins, such as lysozyme and lactoferrin, are part of humoral innate immunity. The aim of this study was to develop a quantitative method, a time-resolved-immunofluorometric assay, to measure lysozyme and lactoferrin in sera, saliva, stools and cervico-vaginal secretions. This method was validated in 51 healthy subjects. Linearity for lysozyme was between 1.02 and 25 microg/l and for lactoferrin between 1.02 and 100 microg/l. The detection limit was 0.5 microg/l for lysozyme and 1 microg/l for lactoferrin. Albumin and alpha1-antitrypsin were measured by immuno-nephelometry to calculate salivary, intestinal and cervico-vaginal coefficients of excretion. Lysozyme and lactoferrin were present in all types of mucosal surfaces. Very high concentrations of lysozyme and lactoferrin were found in cervico-vaginal fluid (166.2 and 72.7 mg/l, respectively), compared to the concentrations found in the other mucosal fluids. Lysozyme in stools was produced at the rate of 0.42 mg/d compared to 0.02 mg/d lactoferrin production. Lysozyme and lactoferrin greatly exceeded the values expected from the molecular weight-affected seepage from plasma, suggesting primarily local synthesis in healthy subjects. Quantitative measurement of lysozyme and lactoferrin can aid in the assessment of the activity of mucus-associated lymphoid tissues in innate immunity, and can help in further understanding of the role of these proteins in mucosal diseases.